20‐kDa chaperone‐like protein of Bacillus thuringiensis ssp. israelensis enhances yield, crystal size and solubility of Cry3A

• Published in: Letters in applied microbiology Vol. 54; no. 2; pp. 88 - 95
• Main Authors: Diaz‐Mendoza, M, Bideshi, D.K, Ortego, F, Farinós, G.P, Federici, B.A
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01.02.2012; Blackwell
• Subjects: 20-kDa protein; Animals; Bacillus thuringiensis; Bacillus thuringiensis - chemistry; Bacillus thuringiensis - genetics; Bacillus thuringiensis - metabolism; Bacillus thuringiensis subsp. israelensis; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacterial Proteins - ultrastructure; Biological and medical sciences; chaperone-like protein; Coleoptera - microbiology; Colorado; Colorado potato beetle; Cry3A; crystallization; crystals; Endotoxins - chemistry; Endotoxins - genetics; Endotoxins - metabolism; fermentation; Fundamental and applied biological sciences. Psychology; genes; Hemolysin Proteins - chemistry; Hemolysin Proteins - genetics; Hemolysin Proteins - metabolism; Hemolysin Proteins - ultrastructure; Larva - microbiology; Leptinotarsa decemlineata; microbial insecticides; Microbiology; Molecular Chaperones - chemistry; Molecular Chaperones - genetics; Molecular Chaperones - metabolism; neonates; Pest Control, Biological; Solubility; spores; Spores, Bacterial - physiology; toxicity; transgenic plants; 20-kDa protein; Chrysomelidae; chaperone-like protein; Coleoptera; Applied microbiology; Insecta; Bacillus thuringiensis; Chaperone; Crystals; Solubility; Bacillaceae; Protein; Bacillales; Leptinotarsa decemlineata; Colorado potato beetle; Arthropoda; Bacteria; Yield; Biopesticide; Invertebrata; Cry3A; Microbial insecticide
• ISSN: 0266-8254; 1472-765X
• DOI: 10.1111/j.1472-765X.2011.03176.x

Aims: To determine whether the 20‐kDa chaperone‐like protein of Bacillus thuringiensis ssp. israelensis enhances synthesis, crystallization and solubility of the Cry3A coleopteran toxin and whether the crystalline inclusions produced are toxic to neonates of the Colorado potato beetle, Leptinotarsa decemlineata. Methods and Results: The cry3A gene was expressed in the 4Q7 strain of B. thuringiensis ssp. israelensis in the absence or presence of the 20‐kDa gene. The 20‐kDa protein enhanced Cry3A yield by 2·7‐fold per unit of fermentation medium. Crystal volumes averaged 2·123 and 0·964 μm3 when synthesized in, respectively, the presence or absence of the 20‐kDa protein. Both crystals were soluble at pH 5 and pH 6; however, the larger crystal was 1·7× and 1·5× more soluble at, respectively, pH 7 and pH 10. No significant difference in toxicity against L. decemlineata neonates was observed. Conclusions: This report demonstrated that the 20‐kDa chaperone‐like protein enhances yield, volume and solubility of the coleopteran Cry3A crystalline inclusions per unit crystal/spore mixture. Significance and Impact of the Study: This is the first report showing that an accessory protein (20‐kDa) could enhance synthesis and crystallization of Cry3A, a finding that could be beneficial for commercial production of this coleopteran‐specific insecticidal protein for microbial insecticides and possibly even for transgenic crops.